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Franck Zal - One of the best experts on this subject based on the ideXlab platform.

  • high level production of recombinant arenicola marina globin chains in escherichia coli a new generation of Blood Substitute
    Artificial Cells Blood Substitutes and Biotechnology, 2009
    Co-Authors: Thomas Harnois, Morgane Rousselot, Helene Rogniaux, Franck Zal
    Abstract:

    This work reports for the first time the expression of a soluble B2 globin chain that is part of the extracellular hexagonal-bilayer haemoglobin from Arenicola marina. Two recombinant B2 globins were produced, one fused with gluthatione S-tranferase (B2-GST) and the other without a fusion tag (RecB2) and requiring a different purification procedure. We also describe a new method for the expression of globin that uses Studier's auto-induction medium together with the heme precursor delta-aminolevulinic acid. Media supplementation with the heme precursor delta-aminolevulinic acid in the culture increased heme synthesis by E. coli leading to the expression of the recombinant B2 globins in their active form. RecB2 and B2-GST were expressed with a yield of up to 105 mg/l of E. coli culture. Our approach is rapid and requires only one chromatographic purification step for B2-GST and three purification steps for RecB2. The overall results on RecB2 and B2-GST show that the recombinant globins exhibit similar properties to those of Arenicola marina native HBL-Hb with a great stability and a strong oxygen binding. The results and methodologies described in this paper are the beginning of a work aiming at reconstituting a recombinant HBL-Hb by genetic engineering in order to produce an innovative oxygen carrier for therapeutic applications.

  • Arenicola marina extracellular hemoglobin: a new promising Blood Substitute
    Biotechnology Journal, 2006
    Co-Authors: Morgane Rousselot, Eric Delpy, Vincent Lagente, Christophe Drieu La Rochelle, Ralph Pirow, Jean-françois Rees, Agnès Hagège, Dominique Le Guen, Stéphane Hourdez, Franck Zal
    Abstract:

    The need to develop a Blood Substitute is now urgent because of the increasing concern over Europe’s BSE outbreak and the worldwide HIV/AIDS epidemic, which have cut Blood supplies. Extracellular soluble hemoglobin has long been studied for its possible use as a safe and effective alternative to Blood transfusion, but this has met with little success. Clinical trials have revealed undesirable side effects–oxidative damage and vasoconstriction–that hamper the application of cellfree hemoglobin as a Blood Substitute. We have addressed these problems and have found a new promising extracellular Blood Substitute: the natural giant extracellular polymeric hemoglobin of the polychaete annelid Arenicola marina. Here we show that it is less likely to cause immunogenic response; its functional and structural properties should prevent the side effects often associated with the administration of extracellular hemoglobin. Moreover, its intrinsic properties are of interest for other therapeutic applications often associated with hemorrhagic shock (ischemia reperfusion, treatment of septic shock and for organ preservation prior to transplantation). Moreover, using natural hemoglobin is particularly useful since recombinant DNA techniques could be used to express the protein in large quantities.

Marcos Intaglietta - One of the best experts on this subject based on the ideXlab platform.

  • deferoxamine lowers tissue damage after 80 exchange transfusion with polymerized hemoglobin
    Antioxidants & Redox Signaling, 2006
    Co-Authors: Pedro Cabrales, Amy G Tsai, Marcos Intaglietta
    Abstract:

    Hemoglobin (Hb) solutions have been proposed as potential Substitutes for erythrocytes to maintain oxygencarrying capacity in situations in which Blood is not available. This study investigated systemic and microvascular hemodynamics as well as tissue oxygenation and viability after an 80% exchange transfusion with an oxygen-carrying Blood Substitute based on polymerized bovine hemoglobin (PBH). Studies were carried in unanesthetized hamsters prepared with a window-chamber model for microcirculation evaluation. Heme iron-mediated injury to the tissue was analyzed by using deferoxamine (an iron chelator), which reduces free iron toxicity. Exchange transfusion led to a significant decrease in hematocrit (Hct) and an increase in plasma Hb, in addition to a significant decrease of arteriolar and venular diameters, flow velocity, and, therefore, microvascular Blood flow. Capillary perfusion was severely compromised after exchange, but tissue pO2 increased above baseline, and oxygen extraction was reduced. Apop...

  • increased tissue po2 and decreased o2 delivery and consumption after 80 exchange transfusion with polymerized hemoglobin
    American Journal of Physiology-heart and Circulatory Physiology, 2004
    Co-Authors: Pedro Cabrales, Amy G Tsai, Marcos Intaglietta
    Abstract:

    The O2-carrying Blood Substitute based on polymerized bovine hemoglobin (PBH) was used to determine efficacy in maintaining tissue Po2 after an 80% isovolemic Blood exchange leading to a hematocrit...

Morgane Rousselot - One of the best experts on this subject based on the ideXlab platform.

  • high level production of recombinant arenicola marina globin chains in escherichia coli a new generation of Blood Substitute
    Artificial Cells Blood Substitutes and Biotechnology, 2009
    Co-Authors: Thomas Harnois, Morgane Rousselot, Helene Rogniaux
    Abstract:

    This work reports for the first time the expression of a soluble B2 globin chain that is part of the extracellular hexagonal-bilayer haemoglobin from Arenicola marina. Two recombinant B2 globins were produced, one fused with gluthatione S-tranferase (B2-GST) and the other without a fusion tag (RecB2) and requiring a different purification procedure. We also describe a new method for the expression of globin that uses Studier's auto-induction medium together with the heme precursor δ-aminolevulinic acid. Media supplementation with the heme precursor δ-aminolevulinic acid in the culture increased heme synthesis by E. coli leading to the expression of the recombinant B2 globins in their active form. RecB2 and B2-GST were expressed with a yield of up to 105 mg/l of E. coli culture. Our approach is rapid and requires only one chromatographic purification step for B2-GST and three purification steps for RecB2. The overall results on RecB2 and B2-GST show that the recombinant globins exhibit similar properties t...

  • high level production of recombinant arenicola marina globin chains in escherichia coli a new generation of Blood Substitute
    Artificial Cells Blood Substitutes and Biotechnology, 2009
    Co-Authors: Thomas Harnois, Morgane Rousselot, Helene Rogniaux, Franck Zal
    Abstract:

    This work reports for the first time the expression of a soluble B2 globin chain that is part of the extracellular hexagonal-bilayer haemoglobin from Arenicola marina. Two recombinant B2 globins were produced, one fused with gluthatione S-tranferase (B2-GST) and the other without a fusion tag (RecB2) and requiring a different purification procedure. We also describe a new method for the expression of globin that uses Studier's auto-induction medium together with the heme precursor delta-aminolevulinic acid. Media supplementation with the heme precursor delta-aminolevulinic acid in the culture increased heme synthesis by E. coli leading to the expression of the recombinant B2 globins in their active form. RecB2 and B2-GST were expressed with a yield of up to 105 mg/l of E. coli culture. Our approach is rapid and requires only one chromatographic purification step for B2-GST and three purification steps for RecB2. The overall results on RecB2 and B2-GST show that the recombinant globins exhibit similar properties to those of Arenicola marina native HBL-Hb with a great stability and a strong oxygen binding. The results and methodologies described in this paper are the beginning of a work aiming at reconstituting a recombinant HBL-Hb by genetic engineering in order to produce an innovative oxygen carrier for therapeutic applications.

  • Arenicola marina extracellular hemoglobin: a new promising Blood Substitute
    Biotechnology Journal, 2006
    Co-Authors: Morgane Rousselot, Eric Delpy, Vincent Lagente, Christophe Drieu La Rochelle, Ralph Pirow, Jean-françois Rees, Agnès Hagège, Dominique Le Guen, Stéphane Hourdez, Franck Zal
    Abstract:

    The need to develop a Blood Substitute is now urgent because of the increasing concern over Europe’s BSE outbreak and the worldwide HIV/AIDS epidemic, which have cut Blood supplies. Extracellular soluble hemoglobin has long been studied for its possible use as a safe and effective alternative to Blood transfusion, but this has met with little success. Clinical trials have revealed undesirable side effects–oxidative damage and vasoconstriction–that hamper the application of cellfree hemoglobin as a Blood Substitute. We have addressed these problems and have found a new promising extracellular Blood Substitute: the natural giant extracellular polymeric hemoglobin of the polychaete annelid Arenicola marina. Here we show that it is less likely to cause immunogenic response; its functional and structural properties should prevent the side effects often associated with the administration of extracellular hemoglobin. Moreover, its intrinsic properties are of interest for other therapeutic applications often associated with hemorrhagic shock (ischemia reperfusion, treatment of septic shock and for organ preservation prior to transplantation). Moreover, using natural hemoglobin is particularly useful since recombinant DNA techniques could be used to express the protein in large quantities.

Pedro Cabrales - One of the best experts on this subject based on the ideXlab platform.

  • deferoxamine lowers tissue damage after 80 exchange transfusion with polymerized hemoglobin
    Antioxidants & Redox Signaling, 2006
    Co-Authors: Pedro Cabrales, Amy G Tsai, Marcos Intaglietta
    Abstract:

    Hemoglobin (Hb) solutions have been proposed as potential Substitutes for erythrocytes to maintain oxygencarrying capacity in situations in which Blood is not available. This study investigated systemic and microvascular hemodynamics as well as tissue oxygenation and viability after an 80% exchange transfusion with an oxygen-carrying Blood Substitute based on polymerized bovine hemoglobin (PBH). Studies were carried in unanesthetized hamsters prepared with a window-chamber model for microcirculation evaluation. Heme iron-mediated injury to the tissue was analyzed by using deferoxamine (an iron chelator), which reduces free iron toxicity. Exchange transfusion led to a significant decrease in hematocrit (Hct) and an increase in plasma Hb, in addition to a significant decrease of arteriolar and venular diameters, flow velocity, and, therefore, microvascular Blood flow. Capillary perfusion was severely compromised after exchange, but tissue pO2 increased above baseline, and oxygen extraction was reduced. Apop...

  • increased tissue po2 and decreased o2 delivery and consumption after 80 exchange transfusion with polymerized hemoglobin
    American Journal of Physiology-heart and Circulatory Physiology, 2004
    Co-Authors: Pedro Cabrales, Amy G Tsai, Marcos Intaglietta
    Abstract:

    The O2-carrying Blood Substitute based on polymerized bovine hemoglobin (PBH) was used to determine efficacy in maintaining tissue Po2 after an 80% isovolemic Blood exchange leading to a hematocrit...

Gerald S. Moss - One of the best experts on this subject based on the ideXlab platform.

  • the first randomized trial of human polymerized hemoglobin as a Blood Substitute in acute trauma and emergent surgery
    Journal of The American College of Surgeons, 1998
    Co-Authors: Steven A. Gould, Ernest E Moore, James B. Haenel, Jon M. Burch, Richard E. Dewoskin, David B Hoyt, Joan Garcia, Gerald S. Moss
    Abstract:

    Abstract Background: Human polymerized hemoglobin (PolyHeme) is a universally compatible, disease-free, oxygen-carrying resuscitative fluid. This is the first prospective, randomized trial to compare directly the therapeutic benefit of PolyHeme with that of allogeneic red Blood cells (RBCs) in the treatment of acute Blood loss. Study Design: Forty-four trauma patients (33 male, 11 female) aged 19–75 years with an average Injury Severity Score (ISS) score of 21 ± 10 were randomized to receive red cells (n = 23) or up to 6 U (300 g) of PolyHeme (n = 21) as their initial Blood replacement after trauma and during emergent operations. Results: There were no serious or unexpected adverse events related to PolyHeme. The PolyHeme infusion of 4.4 ± 2.0 units (mean ± SD) resulted in a plasma [Hb] of 3.9 ± 1.3 g/dL, which accounted for 40% of the total circulating [Hb]. There was no difference in total [Hb] between the groups before infusion (10.4 ± 2.3 g/dL control vs. 9.4 ± 1.9 g/dL experimental). At end-infusion the experimental RBC [Hb] fell to 5.8 ± 2.8 g/dL vs. 10.6 ± 1.8 g/dL (p Conclusions: PolyHeme is safe in acute Blood loss, maintains total [Hb] in lieu of red cells despite the marked fall in RBC [Hb], and reduces the use of allogeneic Blood. PolyHeme appears to be a clinically useful Blood Substitute.

  • Clinical Utility of Human Polymerized Hemoglobin as a Blood Substitute after Acute Trauma and Urgent Surgery
    Journal of Trauma-injury Infection and Critical Care, 1997
    Co-Authors: Steven A. Gould, Ernest E Moore, Frederick A. Moore, James B. Haenel, Jon M. Burch, Hansa L. Sehgal, Lakshman R. Sehgal, Richard E. Dewoskin, Gerald S. Moss
    Abstract:

    We have previously documented the safety of 1 unit (50 gram) of human polymerized hemoglobin (Poly SFH-P) in healthy volunteers. This report describes the first patient trial to assess the therapeutic benefit of Poly SFH-P in acute Blood loss. Thirty-nine patients received 1 In = 14), 2 In = 2), 3 In = 15), or 6 In = 8) units of Poly SFH-P instead of red cells as part of their Blood replacement after trauma and urgent surgery. There were no safety issues related to the infusion of Poly SFH-P. The plasma hemoglobin concentration ([Hb]) after the infusion of 6 units (300 gram) of Poly SFH-P was 4.8 ± 0.8 g/dL (mean ± SD). Although the red cell [Hb] fell to 2.9 ± 1.2 g/dL, the total [Hb] was maintained at 7.5 ± 1.2 g/dL. Poly SFH-P maintained total [Hb], despite the marked fall in red cell [Hb] due to Blood loss. The utilization of O 2 (extraction ratio) was 27 ± 16% from the red cells and 37 ± 13% from the Poly SFH-P. Twenty-three patients (59%) avoided allogeneic transfusions during the first 24 hours after Blood loss. Poly SFH-P effectively loads and unloads O 2 and maintains total hemoglobin in lieu of red cells after acute Blood loss, thereby reducing allogeneic transfusions. Poly SFH-P seems to be a clinically useful Blood Substitute.

  • Clinical Development of Human Polymerized Hemoglobin as a Blood Substitute
    World Journal of Surgery, 1996
    Co-Authors: Steven A. Gould, Gerald S. Moss
    Abstract:

    Although the efficacy of hemoglobin-based oxygen carriers was established more than 60 years ago, all prior clinical trials have demonstrated significant toxicity characterized by renal dysfunction, gastrointestinal distress, and systemic vasoconstriction. The mechanisms of these toxicities now appear to be understood. Tetrameric forms of the hemoglobin molecule extravasate from the circulation and interact with endothelium-derived relaxing factor, leading to unopposed vasoconstriction. Although numerous efforts are under way to chemically modify the native tetramer, it is likely that all tetrameric forms of the hemoglobin molecule will continue to extravasate. We have focused on developing a polymerized form of hemoglobin that is virtually free of unreacted tetramer. The development and characterization of this polymerized pyridoxylated hemoglobin solution (Poly SFH-P) is described. Clinical trials have been completed successfully in volunteers and are now under way to assess the safety and efficacy of Poly SFH-P as a clinically useful red Blood cell Substitute for treatment of acute Blood loss in the setting of trauma and surgery.